Agmatine Is Efficiently Transported by Non-Neuronal Monoamine Transporters Extraneuronal Monoamine Transporter (EMT) and Organic Cation Transporter 2 (OCT2)

Abstract

Agmatine has received considerable attention recently. Available evidence suggests that agmatine functions as a neurotransmitter and inhibits, via induction of antizyme, cellular proliferation. Because of its positive charge, agmatine will not appreciably cross cellular membranes by simple diffusion. Indeed, all physiological models require a channel or transporter protein in the plasma membrane to effect inactivation or nonexocytotic release of agmatine. However, a transport mechanism for agmatine has not been identified on a molecular level so far. In the present study, the non-neuronal monoamine transporters, organic cation transporter (OCT) 1, OCT2, and extraneuronal monoamine transporter (EMT) (gene symbols SLC22A1-A3), both from human and rat, were examined, stably expressed in 293 cells, for [3 H]agmatine transport. Our results indicate that OCT2 and EMT, but not OCT1, efficiently translocate agmatine. The structural homolog putrescine was not accepted as substrate. Uptake of agmatine via EMT and OCT2 was saturable, with K m values of 1 to 2 mM. The affinity of OCT1 was 10-fold lower. Carrier-mediated efflux of agmatine was documented in a trans-stimulation experiment. Finally, uptake of agmatine increased dramatically with increasing pH. Thus, only the singly charged species of agmatine is accepted as substrate. In conclusion, both EMT and OCT2 must be considered for the control of agmatine levels in rat and human.In recent years, agmatine has been recognized as an extracellular signaling substance in mammalia. Agmatine [1-(4-aminobutyl)guanidine] is an amine, synthesized by decarboxylation of L-arginine by arginine decarboxylase. Arginine decarboxylase activity is associated with mitochondrial membranes and is most prevalent in kidney, liver, and brain (Lortie et al., 1996). Agmatine is present in plasma and is widely but unevenly distributed in mammalian tissue (Raasch et al., 1995).There is evidence that agmatine acts as an antiproliferative molecule through induction of the protein antizyme (Satriano et al., 1998;Babal et al., 2001). Antizyme inhibits ornithine decarboxylase and hence polyamine (putrescine, spermine, and spermidine) biosynthesis. At the same time, antizyme suppresses polyamine uptake. Concerted intracellular polyamine depletion eventually leads to growth arrest. Thus, agmatine has been considered a tumor suppressor in the control of cellular proliferation (Satriano et al., 1999). In addition, agmatine might serve as a neurotransmitter or neuromodulator. It is synthesized in specific regions of the brain, stored in synaptic vesicles, released by depolarization, and inactivated by agmatinase and diamine oxidase (Li et al., 1994). Moreover, agmatine binds to ␣ 2 -adrenoceptors and imidazoline binding sites, and blocks NMDA receptor channels and other ligand-gated cation channels. It also inhibits nitric oxide synthase and induces release of peptide hormones. Although the precise function of endogenously released agmatine in the central nervous system is presently still unclear, ...

Biological activity of antitumoural MGBG: the structural variable

“…In addition, FCCP, which collapses the electrochemical gradient of ''in situ'' mitochondria, exhibits a partial inhibition of agmatine transport, as it only excludes the entrance of the amine in the mitochondrial matrix. This data clearly shows that agmatine is transported through an electroneutral mechanism in hepatocyte cultures, supporting the statement that the unprotonated species is involved in this process as observed for EMT and OCT2 transporters (Grundemann et al 2003). Figure 5 also shows that MGBG is transported in hepatocytes, even to a larger extent than agmatine, suggesting that the presence of two guanidinium groups in its molecule further favours this mechanism.…”

“…3), this hypothesis should not be disregarded. Moreover, MGBG could be transported in the unprotonated, uncharged form, since those transporters exhibit an electroneutral behaviour (Grundemann et al 2003).…”

Interaction of Ethambutol with Human Organic Cation Transporters of the SLC22 Family Indicates Potential for Drug-Drug Interactions during Antituberculosis Therapy

“…Tissue culture. Derivation of stably transfected Chinese hamster ovary (CHO) cell lines expressing hOAT1 (CHO-hOAT1) and stably transfected human embryonic kidney 293 (HEK) cells expressing hOAT3 (HEK-hOAT3), hOCT1 (HEK-hOCT1), hOCT2 (HEK-hOCT2), or hOCT3 (HEK-hOCT3), as well as their corresponding empty vectortransfected background control cell lines, has been described previously (16)(17)(18)(19). CHO-hOAT1 cells were maintained at 37°C with 5% CO 2 in phenol red-free RPMI 1640 medium (Gibco-Invitrogen, Grand Island, NY) containing 10% serum, 1% penicillin/streptomycin, and 1 mg/ml G418.…”

Low-affinity uptake of the fluorescent organic cation 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (4-Di-1-ASP) in BeWo cells

“…Not only did TEA and choline fail to inhibit the uptake of 4-Di-1-ASP, but ergothioneine-a specific substrate for OCTN1 [22]-also showed no inhibitory effects ( Figure 5C). Amiloride [6], MPP + [15], and agmatine [23], three other OCT inhibitors, likewise showed no significant inhibition of 4-Di-1-ASP uptake ( Figure 6). …”